The main objectives of this competing continuation application are to elucidate the molecular mechanisms by which the proteasome inhibitor PS-341 induces apoptosis in head and neck squamous cell carcinoma (HNSCC) cells. HNSCC typically presents as a very malignant tumor. Despite significant advances in surgical techniques and chemotherapy for patients with HNSCC, the five year survival rate is the lowest for any major cancer. Therefore, improving the efficacy of cancer therapies has become an urgent topic for both clinical oncologists and basic scientists. Recently, PS-341 has emerged as a novel promising chemotherapeutic agent. Although PS-341 has been found to induce apoptosis in a wide variety of HNSCC cells, the molecular mechanisms by which PS-341 induces apoptosis in these cells are not clear. The preliminary studies presented in this application demonstrate that, in addition to the inhibition of pro-survival transcription factor nuclear factor kappa B (NF-icB), PS-341 potently stimulates the pro-apoptotic ER stress signaling pathway to activate caspase cascade, thereby inducing apoptosis in HNSCC cells. Currently, cisplatin is one of the most common anti-tumor agents for the treatment of HNSCC. Unfortunately, many tumors can gradually develop resistance after initially responding to cisplatin treatment. Importantly, we also observed that PS-341 potently induced apoptosis in cisplatin-resistant SCC cells. Based on these exciting findings, our main hypothesis is that, unlike common chemotherapeutic drugs such as cisplatin, PS-341 may activate a unique apoptotic signaling pathway to induce apoptosis in SCC cells. To further understand the precise mechanism by which PS-341 induces apoptosis in HNSCC cells, four specific aims are proposed. Aim 1 is to explore the role of cytosolic free calcium and identify initiator caspase in PS-341-induced apoptosis in SCC cells. Aim 2 is to explore how PS-341 induces expression of the BH3-only proteins Puma and Noxa and determine whether they play a critical role in PS-341-induced apoptosis. Aim 3 is to explore whether we can potentiate PS-341-mediated apoptosis in SCC cells by inhibiting kB kinase activity induced by PS-341. Aim 4 is to explore whether and how PS-341 induces apoptosis in cisplatin-resistant SCC cells in vitro and in vivo. New findings from this application will have clinical implications for treating HNSCC and may help to develop innovative strategies for human cancer treatment.